The epithelial cell-specific clathrin adaptor AP-1B has a well-established role in polarized sorting of cargos to the basolateral membrane. Here we show that β1 integrin was dependent on AP-1B and its co-adaptor, autosomal recessive hypercholesterolemia protein (ARH), for sorting to the basolateral membrane. We further demonstrate an unprecedented role for AP-1B at the basal plasma membrane during collective cell migration of epithelial sheets. During wound healing, expression of AP-1B (and ARH in AP-1B-positive cells) slowed epithelial-cell migration. We show that AP-1B colocalized with β1 integrin in focal adhesions during cell migration using confocal microscopy and total internal reflection fluorescence (TIRF) microscopy on fixed specimens. Further, AP-1B labeling in cell protrusion was distinct from labeling for the endocytic adaptor complex AP-2. Using stochastic optical reconstruction microscopy (STORM) we identified numerous AP-1B-coated structures at or close to the basal plasma membrane in cell protrusions. In addition, immuno-electron microscopy (EM) showed AP-1B in coated pits and vesicles at the plasma membrane during cell migration. Lastly, qRT-PCR analysis of human epithelial-derived cell lines revealed a loss of AP-1B expression in highly migratory metastatic cancer cells suggesting that AP-1B's novel role at the basal plasma membrane during cell migration might be an anti-cancer mechanism.

上皮细胞特异性网格蛋白衔接子AP-1B在货物到基底外侧膜的极化分选中具有公认的作用。在这里，我们显示β1整联蛋白依赖于AP-1B及其共同适配器常染色体隐性高胆固醇血症蛋白（ARH）来分选到基底外侧膜。我们进一步证明AP-1B在上皮层的集体细胞迁移过程中在基底质膜上的空前作用。在伤口愈合期间，AP-1B（和AP-1B阳性细胞中的ARH）的表达减慢了上皮细胞的迁移。我们显示，在共聚焦显微镜和全内反射荧光（TIRF）显微镜对固定标本进行细胞迁移期间，AP-1B与β1整联蛋白共定位于粘着斑。进一步，细胞突起中的AP-1B标记与内吞衔接子复合物AP-2的标记不同。使用随机光学重建显微镜（STORM），我们在细胞突起的基底质膜处或附近发现了许多AP-1B涂层结构。此外，免疫电子显微镜（EM）显示在细胞迁移过程中，质膜上有涂层的凹坑和囊泡中的AP-1B。最后，对人类上皮细胞系的qRT-PCR分析显示高度迁移的转移性癌细胞中AP-1B表达的丧失，表明AP-1B在细胞迁移过程中在基底质膜上的新作用可能是一种抗癌机制。免疫电子显微镜（EM）显示AP-1B在细胞迁移过程中质膜的包被的凹坑和囊泡中。最后，对人类上皮细胞系的qRT-PCR分析显示高度迁移的转移性癌细胞中AP-1B表达的丧失，表明AP-1B在细胞迁移过程中在基底质膜上的新作用可能是一种抗癌机制。免疫电子显微镜（EM）显示AP-1B在细胞迁移过程中在质膜的包被的凹坑和囊泡中。最后，对人类上皮细胞系的qRT-PCR分析显示高度迁移的转移性癌细胞中AP-1B表达的丧失，表明AP-1B在细胞迁移过程中在基底质膜上的新作用可能是一种抗癌机制。